The present invention relates to a method and apparatus for obtaining improved signal relative to noise without loss of ion collection efficiency for use in mass spectrometry, including liquid chromatography/mass spectrometry, especially as regards to the technique of generating analyte ions known as atmospheric pressure photoionization (APPI).
Liquid chromatography and mass spectrometry have proven powerful analytical tools in identifying molecular components of our world. Liquid chromatography is a fundamental separation technique. Mass spectrometry is a means of identifying xe2x80x9cseparatedxe2x80x9d components according to their characteristic xe2x80x9cweightxe2x80x9d or mass-to-charge ratio. The liquid effluent from liquid chromatography is prepared for ionization and analysis using any of a number of techniques. A conventional technique, atmospheric pressure ionization-electrospray (or simply xe2x80x9celectrosprayxe2x80x9d, for short), involves spraying the sample into fine droplets.
Early systems which employed electrospray liquid chromatography/mass spectrometry techniques utilized flow splitters that divided the high performance liquid chromatography column effluent. As a result of the effluent splitting, only a small portion, typically 5-50 micro liters per minute, was introduced into the xe2x80x9cspray chamberxe2x80x9d. The bulk of the column effluent did not enter the spray chamber, but went directly to a waste or fraction collector. Because electrospray/mass spectrometry generally provides a concentration sensitive detector, it was not necessary to analyze the entire column effluent flow to obtain sensitive results. Results obtained by splitting are comparable in sensitivity to those obtained by introduction of the entire column effluent flow into the spray chamber (assuming equal charging and sampling efficiencies). Such low flow rates enabled generation of an electrosprayed aerosol solely through the manipulation of electrostatic forces. However, the use of flow splitters has performed poorly in that they experience plugging problems and poor reproducibility.
Newer electrospray systems generate a charged or ionized aerosol through the combination of electrostatic forces and some form of assisted nebulization. Nebulization is the process of breaking a stream of liquid into fine droplets. Nebulization may be xe2x80x9cassistedxe2x80x9d by a number of means, including but not limited to pneumatic, ultrasonic or thermal assists. The assisted nebulization generates an aerosol from the high performance liquid chromatography column effluent, while electric fields induce a charge on the aerosol droplets. The charged aerosol undergoes an ion evaporation process whereby desolvated analyte ions are produced. Ideally, only the desolvated ions enter the mass spectrometer for analysis.
It is a desired feature of an assisted nebulizer system that the vacuum system leading to the mass spectrometer permit desolvated ions to enter, but do not permit relatively large solvated droplets present in the electrosprayed aerosol to enter. Several design approaches are currently in use, but none of the assisted nebulization methods currently practiced provide reliable sensitivity along with robust instrumentation.
In conventional electrospray/nebulization mass spectrometry systems, the electrosprayed aerosol exiting from the nebulizer is sprayed directly towards the sampling orifice or other entry into the vacuum system. That is, the electrosprayed aerosol exiting from the nebulizer and entry into the vacuum system are located along a common center axis, with the nebulizer effluent pointing directly at the entry into the vacuum system and with the nebulizer being considered to be located at an angle of zero (0) degrees relative to the common center axis.
One conventional approach directed at improving performance adjusts the aerosol to spray xe2x80x9coff-axisxe2x80x9d. That is, the aerosol is sprayed xe2x80x9coff-axisxe2x80x9d at an angle of as much as 45 degrees with respect to the center axis of the sampling orifice. In addition, a counter current gas is passed around the sampling orifice to blow the solvated droplets away from the orifice. The gas velocities typically used generate a plume of small droplets. Optimal performance appears to be limited to a flow rate of 200 microliters per minute or lower.
In another system, an aerosol is generated pneumatically and aimed directly at the entrance of a heated capillary tube. The heated capillary exits into the vacuum system. Instead of desolvated ions entering the capillary, large charged droplets are drawn into the capillary and the droplets are desolvated while in transit. The evaporation process takes place in the capillary as well. Exiting the capillary in a supersonic jet of vapor, the analyte ions are subsequently focused, mass analyzed and detected.
This system has several disadvantages and limitations, including sample degradation, re-clustering, and loss of sensitivity. Sensitive samples are degraded due to the heat. In the supersonic jet expansion exiting the capillary, the desolvated ions and vapor may recondense, resulting in solvent clusters and background signals. While these clusters may be re-dissociated by collisionally induced processes, this may interfere in identification of structural characteristics of the analyte samples. The large amount of solvent vapor, ions and droplets exiting the capillary require that the detector be arranged substantially off-axis with respect to the capillary to avoid noise due to neutral droplets striking the detector. Removing the large volume of solvent entering the vacuum system requires higher capacity pumps.
Still another conventional system generates the electrosprayed aerosol ultrasonically, uses a counter current drying gas, and most typically operates with the electrosprayed aerosol directed at the sampling capillary. One disadvantage of this configuration is that optimal performance is effectively limited to less than five hundred (500) microliters per minute. Adequate handling of the aqueous mobile phase is problematic. Furthermore, the apparatus is complex and prone to mechanical and electronic failures.
In another conventional system, a pneumatic nebulizer is used at substantially higher inlet pressures (as compared with other systems). This results in a highly collimated and directed electrosprayed aerosol. This aerosol is aimed off axis to the side of the orifice and at the nozzle cap. Although this works competitively, there is still some noise which is probably due to stray droplets. The aerosol exiting the nebulizer has to be aimed carefully to minimize noise while maintaining signal intensity. Thus, repeated and tedious adjustments are often required.
In addition to atmospheric pressure ionization-electrospray, another conventional technique for preparing a liquid effluent for ionization and analysis is atmospheric pressure chemical ionization. Fundamentally, atmospheric pressure chemical ionization involves the conversion of the mobile phase and analyte from the liquid to the gas phase and then the ionization of the mobile phase and analyte molecules. Atmospheric pressure chemical ionization is a soft ionization technique that yields charged molecular ions and adduct ions. Atmospheric pressure chemical ionization actually includes several distinct ionization processes, with the relative influence of each process dependent on the chemistry of the mobile phase and the analyte.
Each of techniques of atmospheric pressure ionization-electrospray and atmospheric pressure chemical ionization is suited to different, and complementary, classes of molecular species. Briefly, atmospheric pressure ionization-electrospray is generally concentration dependent (that is to say, higher concentration equals better performance), and performs well in the analysis of moderately to highly polar molecules. It works well for large, biological molecules and pharmaceuticals, especially molecules that ionize in solution and exhibit multiple charging. Atmospheric pressure ionization-electrospray also performs well for small molecules, provided the molecule is fairly polar. Low flow rates enhance the performance of the atmospheric pressure ionization-electrospray technique. Atmospheric pressure chemical ionization, on the other hand, performs with less dependence on concentration and performs better on smaller non-polar to moderately polar molecules. Higher flow rates enhance the performance of the atmospheric pressure chemical ionization technique. However, there are still analytes that do not ionize at all when these ionization techniques are employed, or which ionize weakly when these ionization techniques are employed.
In addition to the two conventionally employed ionization techniques of atmospheric pressure ionization-electrospray and atmospheric pressure chemical ionization, an alternative technique which has been developed for producing ions from a liquid sample is referred to as atmospheric pressure photoionization (APPI). Generally, the technique of atmospheric pressure photoionization provides a method of analyzing a sample of an analyte provided as a sample solution. According to one such technique, the sample solution is formed into an aerosol spray, for example in a nebulizer, and the solvent is evaporated. The sample stream is irradiated, e.g., subjected to photons, in a region at atmospheric pressure, in the vapor state after evaporation of the sprayed droplet. Collisions between the photons and the analyte result in ionization of the analyte. The analyte ions are passed from the atmospheric pressure ionization region into a mass analyzer for mass analysis.
According to another such technique, dopant is provided, either separately or as the solvent of the sample solution. The sample solution is formed into a spray, for example in a nebulizer, and the solvent is evaporated. The sample stream is irradiated, e.g., subjected to photons, in a region at atmospheric pressure to ionize the dopant. Again, this irradiation step takes place when the sample is in the vapor state after evaporation of the sprayed droplet. Then subsequent collisions between the ionized dopant and the analyte result in ionization of the analyte. Analyte ions are passed from the atmospheric pressure ionization region into a mass analyzer for mass analysis. This technique has been found to give enhanced ionization for some substances, as compared to atmospheric pressure chemical ionization.
Configurations for APPI in present use often provide unsatisfactory signal relative to noise and do not provide for optimal ion collection efficiency. Therefore, there exists a need for an improved method and apparatus for obtaining improved signal relative to noise without loss of ion collection efficiency for use in mass spectrometry, including liquid chromatography/mass spectrometry, especially as regards the technique of generating analyte ions known as atmospheric pressure photoionization.
The invention comprises an atmospheric pressure ion source, e.g. for a mass spectrometer, that produces ions by atmospheric pressure photoionization (APPI). It includes a vaporizer, a photon source for photoionizing vapor molecules upon exit from the vaporizer, a passageway for transporting ions to, for example, a mass spectrometer system, and a means for directing the ions into the passageway. In one embodiment, the passageway has a center axis situated substantially orthogonal to the center axis of the vaporizer. In another embodiment, the center axis of the passageway and the center axis of the vaporizer define an angle in the range of about 20 degrees to 180 degrees.
Included in the invention is a method for creating and transporting ions in an atmospheric pressure ion source by forming them with atmospheric pressure photoionization along an axis and directing them into a passageway oriented at an angle to that axis.